Experiments are proposed to develop mechanism-based, energy restriction-driven interventions that promote the elimination of transformed cells from a tissue by apoptosis. This work is based on our current effort to identify the mechanisms that explain how limiting energy availability by dietary energy restriction (DER) or energy restriction mimetic agents (ERMA) inhibits mammary carcinogenesis and how some clones of transformed cells evade protective activity. The mechanisms that will be investigated include inhibition of autophagy, blocking the switch to aerobic glycolysis, and inhibition of sirtuin-mediated deacetylation. Proof- in-principle experiments are proposed to: 1) determine if chloroquine, an inhibitor of autophagy, enhances the cancer inhibitory activity of energy restriction. We hypothesize that induction of autophagy by energy restriction decreases the cancer inhibitory activity of these interventions by blunting the induction of apoptosis in favor of enhancing cell survival/stress resistance mechanisms; 2) investigate whether dichloroacetate, a drug that reverses the switch to aerobic glycolysis observed in cancers (the Warburg effect), can be used to prevent the development of breast cancer and to increase the cancer inhibitory activity of energy restriction. We hypothesize that reversal of aerobic glycolysis will inhibit mammary carcinogenesis and enable energy restriction mediated apoptotic death of transformed cells; and 3) evaluate how the induction of SIRT1 by energy restriction alters the effectiveness of energy restriction in inhibiting mammary carcinogenesis. Our hypothesis is that SIRT1, a class III histone deacetylase that is induced by energy restriction, blunts cancer inhibition mediated by AMPK activation and AKT down regulation. A rapid emergence model for breast cancer will be used to conduct these experiments. When required by the questions being addressed, epithelial cells will be excised from mammary gland and mammary carcinomas using laser capture microdissection. Methods will include transmission electron microscopy, RT-PCR, western blots, reverse phase protein arrays, IHC, and activity assays using fluorescent polarization detection systems. This work has the potential to provide additional new insights about the mechanisms by which energy restriction inhibits mammary carcinogenesis and to define interventions that promote the selective deletion of transformed cells from the breast. If this potential is realized, it would represent a major advance in cancer prevention research and may have applicability to the control of cancer at organ sites in addition to the breast.